Hygiena MicroSnap TOTAL Instruction Sheet

1 INS0126 JUNE 2012 Rev. A Direction for use of MicroSnap TOTAL (Total Microbial Activity Test) Part#: ATP-ENT200 & ATP-APC200 Log RLU Parts Required: 1. MicroSnap TOTAL Enrichment Swab Device, Part # ATP-ENT200 (for environmental surfaces, liquids and food suspensions) 2. MicroSnap TOTAL Detection Device, Part #ATP-APC200 Description/ Intended Use: MicroSnap TOTAL (Total Microbial Activity Test) is a rapid bioluminogenic test method for the detection and enumeration of total population of bacteria or total contamination in a sample providing results in less than 8 hours. MicroSnap TOTAL consists of an Enrichment Swab Device containing proprietary growth medium and a Detection Device containing bioluminogenic reagents in which biomarkers produced by bacteria are measured using a small portable luminometer. The two step test procedure requires a short incubation period to enhance the growth of bacteria followed by the detection step. During incubation in the Enrichment broth, the number of bacteria is increased and potential sample interference is reduced. As the bacteria grow they use up their available food resources in the broth and generate enzymatic biomarkers, such that the greater the number of bacteria, the higher the biomarker concentration after a set time interval. Subsequently, the biomarker produced by living and growing bacteria is measured using a luminometer. The light output is directly proportional to the concentration of biomarkers and number of bacteria present.[ A relationship between biomarker concentration at a specific point and starting numbers of mixed bacteria can be established.] MicroSnap TOTAL can be used to test environmental surfaces, foodstuff, water and other filterable liquids. The test should be run using aseptic techniques in a laboratory or in a controlled environment.. Material and Reagents required but not provided: Sample preparation diluents and equipment Recommended diluents for product samples: o Buffered Peptone Water o Maximum Recovery Diluent o Butterfields o Other validated diluents of users’ choice Recommended Equipment: o Incubator set at 37°C ± 1°C (Hygiena optional) o Luminometer ? EnSURE, Part#: ATP-205, or ? SystemSURE Plus, Part#: ATP-201 Test Procedure: Refer to diagrams in the following pages. Step 1: Enrichment For environmental surfaces, liquids and solid foodstuffs A. Collect sample and place in the MicroSnap TOTAL Enrichment Swab (Part. # ATP-ENT200) Samples can be of the following types; i. Surface Swabs (typically 4 x 4 inches; 10 x 10 cm) see diagram Step 1 A1. ii. 1mL liquid food, beverage or water samples added directly to Enrichment Swab. Step 1 A2. iii. 1mL 10% w/v food homogenate added directly to Enrichment Swab. The food homogenate is prepared using recommended diluents and standard microbiological procedures. Step 1 A3. B. Re-insert the swab into swab tube. Step 1 B. C. Activate device by bending the bulb to break the Snap Valve pin at the base. Step 1 C. D. After the Snap Valve pin is broken, separate the bulb / swab assembly (about 1 – 2”) from the swab tube to release the internal pressure and squeeze the bulb 1 or 2 times to release the Enrichment Broth into swab tube. Most of the Enrichment Broth will now be at the bottom of the swab tube. Reinsert the bulb / swab assembly firmly into the swab tube to seal the device. Step 1 D. E. Shake the tube gently to mix sample and Enrichment Broth. Step 1 E. F. Incubate at 37° ± 1°C for 7 hours. Step 1 F. This will be the Enriched Sample. Step 2: Detection A. Allow the MicroSnap TOTAL Detection Device (ATP-APC200) to equilibrate to room temperature (10 minutes). Shake the test device by tapping on the palm of your hand 5 times (to bring the droplets of liquid dispersed in the tube to the bottom of the tube; prior to adding the Enriched Sample to the tube. This will facilitate the mixing of the Enriched Sample with the solution in the tube. Step 2 A. B. Aseptically remove an aliquot of the sample from the Enrichment Swab (ATP-ENT200) and transfer it to the MicroSnap TOTAL Detection Device (ATP-APC200). Optimum volume is 0.1ml, (or 2-3 drops). Filling to the marked fill line is exactly 0.1ml. Excess fill does not affect the test. A simple and easy way to transfer sample from the Enrichment Swab to the MicroSnap TOTAL detection device is to squeeze the bulb of the Enrichment Swab. This will suck the sample back up into the Snap Valve bulb. Then use the upright, press “OK” button to initiate the measurement. Results will appear after the15 second count down. Step 2 E. F. Read result as RLU from the display on luminometer and interpret the result as directed below. Step 2 F. Interpretation of Results: The results on the luminometer are displayed as Relative Light Units (RLU). The RLU output is proportional to the starting inoculums and the corresponding bacteria equivalent numbers (expressed as Colony Forming Units, CFU). The Table 1 below shows the equivalent CFU values for RLU measurements obtained using the 2 different Hygiena luminometers. Table 1 Correlation between CFU/g or mL and average RLU After 7 hour Incubation at 37°C CFU/ml or g SystemSURE Plus EnSURE 0 = 10 = 20 1-9 11-20 21-40 10-50 21-60 41-120 51-100 61-120 121-240 101-1000 121-1200 241-2400 1001-10000 1201-9000 2401-9000 Typical Correlation of Dilutions of Food After 7 Hour Incubation 10000 1000 100 10 1 1 10 100 1000 10000 Log CFU/mL Controls It is advisable to run positive and negative controls according to Good Laboratory Practices. Safety & Precautions: Components of MicroSnap TOTAL (ATP-ENT200 & ATP-APC200) devices do not pose any health risk when used correctly. Used devices that confirm positive results may be bio- hazardous and should be disposed of safely in compliance with Good Laboratory Practice and Health and Safety Regulations. 1. ATP-APC200 Device is designed for a single use. Do not reuse. 2. Do not use devices after Expiration Date. 3. Sampling should be done aseptically, to avoid cross contamination. 4. Ensure proper incubation temperature and time for the test application. 5. When activating devices, ensure that all the liquid in the bulb is transferred to the tube below. Storage & Shelf Life: Boxes and bags of devices must be stored at 2°-8°C. Devices have a shelf life of 12 months. Check expiration date on label. Caution and User Responsibility: 1. As with any culture medium, MicroSnap TOTAL results do not constitute a guarantee of quality of food, beverage products or processes that are tested with these devices. 2. The user must train personnel in proper testing techniques. Enrichment Swab as a Pasteur pipette for convenience. Remove the bulb from the tube and carefully dispense 2-3 drops (~ 0.1ml) into the Detection Device (ATP-APC200). Step 2 B. Re-insert Snap valve bulb, push all the way in to seat in tube. C. Activate Detection Device (ATP-APC200). Bend the bulb to break the Snap Valve pin. Squeeze bulb 3 times to release the reagent. Step 2 C. D. Shake gently to mix. Step 2 D. E. Insert the whole device into the luminometer, close the lid and holding the unit 2 INS0126 JUNE 2012 Rev. A Direction for use of MicroSnap TOTAL (Total Microbial Activity Test) Part#: ATP-ENT200 & ATP-APC200 MicroSnapTM Total Step: 1 Enrichment of Environmental Surface Swab, Liquid and Solid Samples Surface A Liquids Solid Samples B OR OR A1. Surface: Swab a 10x10cm area or larger depending on protocol with the MicroSnap TOTAL Enrichment Swab (ATP-ENT200). C A2. Liquids: 1ml liquid food, beverage or water sample added directly to MicroSnap TOTAL Enrichment Swab (ATP-ENT200). D A3. Solid Samples: 1ml 10% w/v suspension of solid samples added directly to MicroSnap TOTAL Enrichment Swab (ATP-ENT200). E B. Re-insert Snap-Valve bulb into swab tube. F C. Activate the device. Bend bulb, snapping the Snap-Valve pin. D. Lift the bulb up (about 1 – 2 inches) and squeeze the bulb to release the liquid into tube. Release pressure from the bulb (the bulb is like a dropper bulb) and replace bulb into the tube. Most liquid should be in the bottom of the tube. E. Shake the tube gently to mix sample in the liquid. F. Incubate at 37° ± 1°C for 7 hours. This is the Enriched Sample. Proceed to Step 2. Step: 2 Detection / Measurement A B C D Allow MicroSnap TOTAL Detection Device, ATP-APC200 to equilibrate to room temperature. Shake tube by tapping on the palm of your hand 5 times to bring liquid in the tube to the bottom of the tube. E B. Aseptically transfer 0.1mL (2 to 3 drops or to fill line) of Enriched Sample from MicroSnap TOTAL Enrichment Swab (ATP-ENT200) into MicroSnap TOTAL Detection Device (ATP-APC200). F C. Activate MicroSnap TOTAL Detection Device (ATP-APC200) by breaking the Snap Valve pin with a snap and squeeze action. Squeeze the bulb to release the liquid into the tube. Liquid should be in the bottom of the tube. D. Shake the tube gently to mix sample in the liquid. Test should be ready within 45 seconds after activation. E. Insert MicroSnap TOTAL Test (ATP-APC200), into a luminometer and initiate the measurement. F. Record the results as RLUs and refer to Table 1 above to interpret the results.